The success of protein microarrays in drug discovery, diagnostics, and other biological applications has been hindered by the reliance on techniques that have been widely used in DNA microarrays. The transition to protein microarrays requires a different approach to array fabrication, as well as target immobilization and characterization. The development of effective and robust methods for protein micro-array immobilization is critical to the application of array technology. This is especially challenging with the wide variety of substrates that are used, such as gold, glass, or plastics. Immobilizing ligands in significant quantities, while retaining functionality and in a cost-effective and timely manner, is the primary objective of array technologies.
Pin printing, the most commonly method used for array fabrication, has an established infrastructure that includes robotics and microtiter plates, which make it an intriguing option for protein microarrays; however, pin printing devices are limited by a number of obstacles that are difficult and time consuming to overcome. The challenges include the optimization of a large number of parameters such as humidity, temperature, and surface energy while combating variable spot morphology caused by surface imperfections and over loading. Highly concentrated samples are often required to generate acceptable microarrays, which in the case of most proteins is inconvenient and cost prohibitive. A promising alternative approach to pin-spotting is the use of continuous flow microfluidics, which provides the ability to deliver ligand samples across a well-defined deposition zone.
The first continuous-flow microfluidic devices utilized for patterning surfaces with specific biomolecules and chemistries were developed for optical detection platforms such as SPR. More complex, arrayed independent microfluidic deposition devices followed later. The advantage of these 2-D microfluidic systems was isolated flow cells depositing biomolecules to specific miniature regions of the surface. Dense microarrays could be created for further use in bioassays by lifting the flow cell, rotating it 90°, and flowing the analyte solution back across the patterned lanes. However, such techniques still do not confine deposition to specific locations to minimize sample depletion as found in pin-printed arrays. As such, improvements to existing spotting techniques continue to be sought through ongoing research and development.